Our goals are to understand the molecular nature of the mechanisms underlying germ cell determination and the establishment of the anterioposterior axis in early Drosophila embryos. The initial strategy has been to use genetic techniques to identify and characterize loci required for germ cell formation. One such locus, tudor, has been identified genetically and the corresponding DNA sequence has been cloned. Immunocytochemical studies using antibodies raised against bacterially synthesized tudor protein have demonstrated that this gene product is predominately localized at the posterior end of the developing oocyte. By 0.25 hrs., the gene product is sequestered in nuclei and mitochrondia, accumulating particularly in the mitochrondia of the posterior pole during the second hour of development (Bardsley et al., 1993). Later, the tudor antigen is found within embryonic nuclei but only in the mitochrondia of the presumptive germ cells. We are now testing the hypothesis that the nuclear form of the protein is essential for segmentation of the embryonic abdomen and that the mitochrondial form is necessary for germ cell determination. Molecular modification of the tudor gene will be used to test this hypothesis and to evaluate the portions of the gene product that are essential for its localization. Mutants with aberrant localization are being analyzed to study the role that localization plays in tudor function during embryogenesis. We are also refining techniques for fast-freezing and freeze substitution ovaries of Drosophila so that we may localize tudor gene products in developing oocytes.